PROJECT SUMMARY/ABSTRACT Cells exchanged between a mother and fetus during pregnancy are now known to establish long-term residence in the other individual creating a legacy of maternal cells in her progeny and cells of fetal origin in women who have been pregnant. The capacity for these naturally acquired cells to confer health benefits is supported by experimental and human studies. However detrimental effects on health may also sometimes occur. Transmissible cancer has been reported in dogs, Tasmanian devils, and in case reports in humans, primarily as mother to offspring transfer. To our knowledge no prior study has asked or addressed the possibility that naturally acquired allogeneic cells could sometimes be the source of a malignancy. Asking when this might occur led to this proposal. Patients who undergo organ transplantation are subject to long-term chronic immune suppression. Organ transplant recipients (OTR) are afflicted by multiple, frequently arising, squamous cell keratinocyte carcinomas (SCC). Recurrence rates and mortality related to SCC are significantly increased compared to non-immunosuppressed individuals. The purpose of this R03 application is to investigate SCC in OTR for allogeneic DNA and cells. Aim 1 will begin by interrogating DNA extracted from OTR SCCs by quantitative PCR (qPCR) for a Y-chromosome specific sequence; disproportionate absence of male DNA in tumors from males or presence of male DNA in tumors from females will be considered potential evidence for allogeneic cancer. Cellular studies will next be done by combined fluorescence in situ hybridization (FISH) with Y- and X-chromosome specific probes and concomitant immunohistochemistry (IHC) to identify, quantify, and phenotype sex-mismatched cells in tumor specimens. Aim 2 will investigate OTR SCCs to determine the specific origin of allogeneic DNA and cells detected in Aim 1. In part 1 of Aim 2 family members will be recruited so HLA and other polymorphism genotyping can be conducted. Results will be evaluated to identify a non-shared HLA (or other) polymorphism unique to the family member. DNA extracted from the tumor specimen will be quantitatively interrogated for the non-shared polymorphism selecting from a panel of HLA- and other polymorphism-specific qPCR assays we have developed for this purpose. qPCR testing will include an assay to detect donor origin DNA in tumors. For patients for whom family members are not available, Aim 2 part 2 will conduct SNP arrays and analysis of paired tumor and peripheral blood samples to determine whether tumor DNA is autochthonous or allogeneic in origin.